This project examines mitochondrial functioning in old age and in pathological states in which decreased energy transduction by mitochondria may compromise tissue survival. Experimental results this year derive from two paradigms relating to heart failure and death of cardiac myocytes. (1) We have investigated a changed intramitochondrial free Ca2+ ([Ca2+]m) as a possible mechanism of the less active pyruvate dehydrogenase which we identified in 2 strains of hamster with an inborn cardiomyopathy (B10 14.6 and TO.2) and described in last year's report. Work with isolated mitochondria showed similar gradients of Ca2+ across the mitochondrial membrane in myopathic and healthy strains - i.e. no intrinsic change in the mitochondrial Ca2+-transport proteins per se. However, measurements of [Ca2+]m in situ in isolated cardiac myocytes from myopathic animals showed lower values compared to healthy animals, thought to be due to lower transients in cytosol [Ca2+] on electrical excitation. These results provide a possible mechanism for the impaired dehydrogenase regulation in this animal. (2) Mitochondrial function has been evaluated in isolated cardiac myocytes using the newly-described fluorescent dye JC- 1, which reports on mitochondrial membrane potential ( delta psi). De- energization of the cells due to anoxia or respiratory chain inhibition allows some remaining measure of contractile function and preservation of delta psi. Additional inhibition of glycolysis leads to collapse of both parameters. Evidently ATP derived from glycolysis can sustain delta psi, under the experimental conditions employed, through a reversal of the mitochondrial protontranslocating ATPase.